Understanding the spatial and temporal variation of nutrient concentrations, loads, and their distribution from upstream tributaries is important for the management of large lakes and reservoirs. The Three Gorges Dam was built on the Yangtze River in China, the world’s third longest river, and impounded the famous Three Gorges Reservoir (TGR). In this study, we analyzed total nitrogen (TN) concentrations and inflow data from 2003 till 2010 for the main upstream tributaries of the TGR that contribute about 82% of the TGR’s total inflow. We used time series analysis for seasonal decomposition of TN concentrations and used non-parametric statistical tests (Kruskal-Walli H, Mann-Whitney U) as well as base flow segmentation to analyze significant spatial and temporal patterns of TN pollution input into the TGR. Our results show that TN concentrations had significant spatial heterogeneity across the study area (Tuo River> Yangtze River> Wu River> Min River> Jialing River>Jinsha River). Furthermore, we derived apparent seasonal changes in three out of five upstream tributaries of the TGR rivers (Kruskal-Walli H ρ = 0.009, 0.030 and 0.029 for Tuo River, Jinsha River and Min River in sequence). ... mehrTN pollution from non-point sources in the upstream tributaries accounted for 68.9% of the total TN input into the TGR. Non-point source pollution of TN revealed increasing trends for 4 out of five upstream tributaries of the TGR. Land use/cover and soil type were identified as the dominant driving factors for the spatial distribution of TN. Intensifying agriculture and increasing urbanization in the upstream catchments of the TGR were the main driving factors for non-point source pollution of TN increase from 2003 till 2010. Land use and land cover management as well as chemical fertilizer use restriction were needed to overcome the threats of increasing TN pollution.